As the main antigen-presenting cell of the skin, Langerhans cells (LC) play a key role in many epidermal phenomena such as contact allergic dermatitis and in host defense mechanisms against skin cancer and bacterial, fungal, and viral infections. However, current investigations into LC biology and function are limited due to the following: l) LC do not proliferate in culture and 2) under standard culture conditions, LC take on an activated or primed phenotype as if antigen had been presented and processed. Thus, the ability to study LC in their basal, unprimed state is severely compromised. In addition, the incorporation of LC into in vitro epidermal tissue models, which would allow the study of the immunological reactions of the skin from a tissue perspective, has not been possible. In order to overcome these problems, this project will develop immortalized, human-derived LC lines. Phase I will concentrate on the establishment and characterization of immortalized LC cell lines. Normal LC will be harvested from human epidermis and immortalized using 3 different methods. The resultant cell lines will be characterized in terms of growth rates, morphology, surface phenotype, and ultrastructure. In addition, the ability of the cell lines to stimulate allogeneic T-cells will be determined. During Phase II, the utility of these cell lines to study contact allergic dermatitis, chemokine receptors, and LC migration will be demonstrated and the optimal cell lines will be incorporated into a three-dimensional epidermal skin model. PROPOSED COMMERCIAL APPLICATIONS: The proposed research will produce stable LC lines which will ultimately allow a greater understanding of the immunological reactions of the skin. The LC lines produced will provide researchers the means by which the biology and function of LC in their native, unprimed state can be studied. Finally, researchers will gain a valuable immunological tool in studying and developing therapies against skin cancer and bacterial, fungal, and viral infections including HIV.